ProjectSummary Mylabisinterestedinthemechanismsthatmaintaintissuehomeostasis,whichiscriticalforproperorgan functionthroughoutourlifetimes.Mutationsarethoughttodisruptcellbehaviorsandhomeostasis,andthuslead todisease.However,recentworkdemonstratedthatagedbutphenotypicallynormaltissues,includingskinblood and intestine, are a mosaic of distinct wild-type and mutant clones. Strikingly, 20-30% of cells bear cancer- associated mutations. Intriguingly, we and others have shown that cells containing just one of these cancer- associatedmutationsoutcompeteneighboringwild-typecellsintheskin.Themechanismsthattoleratebutalso restrictmultiplemutantcloneswithininagedtissuesarenotknown. Wehypothesizethattheaccumulationofdiversemutantclonespromoteshealthyaging,aslongastheir clonal outgrowths are confined, and homeostasis is maintained. Mutations have long been equated with the emergenceofpathologyandthereforeasdeleteriousalterationsthatmustbeeradicated.Thehighfrequencyof mutantcellswithinournormal,agedtissueswouldimplyacontinuousandenergeticallyconsuminginvestment tocountertheirputativenegativeconsequence.Alternatively,mutantcells,particularlythosecarryingmutations thatenhance proliferation/growth, might support or evenhelp tissues maintainhomeostasis duringaging. Our unconventional hypothesis predicts that tissues benefit from the increased growth introduced by low- levelgenetic heterogeneity. We propose that a balance is achieved by activation of a mechanism that suppresses aberrant expansion but toleratesand positively utilizesmutant subpopulations.In this scenario, diseasearisesonlyafterathresholdfortoleranceisexceeded,eitherduetolossoftheprotectivemechanisms orduetoanexcessivemutationalburden. InthisPioneerproposal,wewillcombineouruniqueabilitytocapturebehaviorsinanintactmammal,to nowdefinethemolecularandmetabolicconsequencesofmutationsandaging.Understandinghowcellsevolve theirgeneexpressionandmetabolicactivitiesinthepresenceofaccumulatingmutationsandastissuesagewill providefundamentalinsightsintohoworgansadaptandremainfunctionalthroughoutourlifetime. Completion oftheproposedworkwillrevealthemechanismsagingskinusestoconstrainmutantsubpopulationsandhow acquiredmutationsinturnimpacttheagingofskin.Thesefindingscouldtransformourstrategiestotreatcancer, which are currently aimed at eliminating all mutant cells ? which we predict would have unintended, adverse outcomes. Theproposed work will shed light notonly ontissuehomeostasis but alsoon theproblem ofaging, whichisthemajorriskfactorfornearlyeverychronicdisease.